Detection of Ecto and Endoparasites Among Wild Rats from Urban Area in Blimbing, Malang, East Java

ACTA VETERINARIA INDONESIANA Special Issues: 95-101, May 2021 P-ISSN 2337-3202, E-ISSN 2337-4373

Research Preliminary Study: Detection of Ecto and Endoparasites Among Wild Rats From Urban Area in Blimbing, Malang, East Java

Shelly Kusumarini R1, Reza Yesica1, Ida Bagus Gde Rama Wisesa1, Jenny Hermanto2, Yustia Nurholizah2, Maria Widyaneni Trinastuti2

1Department of Parasitology, Faculty of Veterinary Medicine, Universitas Brawijaya 2Student on Faculty of Veterinary Medicine, Universitas Brawijaya *Corresponding author: [email protected] Submitted March 26, 2021 Accepted May 8, 2021

ABSTRACT Zoonotic parasitic infections is one of the global public health issues. The complex parasite transmission allows for the relationship between people, animals, vectors, and the environment. The existence of rat in the environment has an important role as a host and reservoir for various types of ecto and endoparasites. This study was conducted to collect informative data on the parasitic infection of wild rats in Blimbing sub-district, Malang city, East Java. A total number of eight wild rat were captured using live-traps from landfills during 4 days. They were classified by sex, weight and body length. The rats were anesthetized, collecting for any ectoparasites and then their carcasses were dissected for examinations of endoparasites. The result of this study show succesfull live-trap of rodents including Rattus norvegicus (87.5%) and Suncus murinus (12.5%). 50% of sampled rodents were male, 37.5% were female, and 12.5% of female the musk shrew. The presence of helminthes infection in all wild rats, namely Hymenolopis nana, Syphacia obvelata, Nippostrongylus brasiliensis. Furthermore, the ectoparasites identification were Xenopsylla cheopis, Echinolaelap echidninus, and blood protozoa identifications Trypanosoma lewisi. Based on the results show 100% of wild rats positive infections of endo-ectoparasites. This study indicates to continuous study among rodents’ parasites in wild rats in different urban areas and analysis of their potential impact on public health. Keywords: public health, landfill, parasites, rat, zoonosis

ABSTRAK Infeksi parasit zoonosis merupakan salah satu masalah kesehatan masyarakat yang terjadi secara global. Transmisi parasit yang kompleks memungkinkan terdapat hubungan antara manusia, hewan, vektor, dan lingkungan. Keberadaan tikus di lingkungan memiliki peran penting sebagai inang dan reservoir untuk berbagai jenis ekto dan endoparasit. Penelitian ini dilakukan untuk mengumpulkan data informatif terkait infeksi parasit pada tikus liar di Kecamatan Blimbing, Kota Malang, Jawa Timur. Sebanyak delapan tikus liar ditangkap menggunakan perangkap dari tempat pembuangan sampah selama 4 hari. Tikus dikelompokkan berdasarkan jenis kelamin, berat badan, dan panjang tubuh. Tikus tersebut dibius, diperiksa dan dikumpulkan ektoparasitnya dan kemudian dibedah untuk pemeriksaan endoparasit. Hasil penelitian ini menunjukkan jenis tikus yang tertangkap adalah Rattus norvegicus (87,5%) dan Suncus murinus (12,5%). Sebanyak 50% adalah tikus jantan, 37,5% tikus betina, dan 12,5% adalah betina dari Suncus murinus. Ditemukan adanya infeksi cacing pada semua tikus yaitu Hymenolopis nana, Syphacia obvelata, Nippostrongylus brasiliensis. Selanjutnya, ektoparasit yang ditemukan diantaranya Xenopsylla cheopis, Echinolaelap echidninus, dan protozoa darah Trypanosoma lewisi. Berdasarkan hasil penelitian ini menunjukkan bahwa 100% tikus liar positif infeksi endo-ektoparasit. Penelitian ini mengindikasikan perlunya studi lebih lanjut mengenai keberadaan parasit pada tikus liar di wilayah perkotaan dan menganalisis potensi bahayanya terhadap kesehatan masyarakat. Kata kunci: kesehatan masyarakat, TPA, parasit, tikus, zoonosis

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INTRODUCTION Objective Zoonotic parasitic infections are one of the This study aimed to undertake the diversity of global public health issues. Now, a major concern rodents parasites and collect informative data on is the potential of zoonotic disease connecting the parasitic infection at the landfill area in Blimbing with wild rodents to spread of most emerging sub-district, Malang city, Indonesia. infectious disease. In fact, Urban ecosystems present various species of rodents (e.g., Rattus norvegicus, Rattus r. diardii, and Rattus norvegicus MATERIAL AND METHODS (Dina & Ustiawan, 2013; Paramasvaran et al., 2009). The existence of wild rodents in the Ethical Statement environment has an important role as hosts of This study was approved by the Institutional ectoparasites as well as reservoirs for various Animal Care and Use Committee of the Brawijaya types of viruses, bacteria, rickettsia, and University, Malang (Ref. No: 074-KEP-UB-2020). All endoparasites such as protozoa, and helminths protocols and procedures for trapping and handling (Moravvej et al., 2015; Tijjani et al., 2020a). of animals in this study are accordance with the The Complex parasite transmission allows for the direction of the Ethics Committee, Brawijaya relationship between people, animals, vectors, and University. the environment. Until now, increasing cases related parasitic zoonosis were reported around the Trapping Locations and Sampling world (WHO, 2019). This condition associated with the ecological system formed dynamically between Trapping of rodents was conducted in the landfill rodent reservoirs populations and humans causing a area in Blimbing sub-district from 10 August to 14 risk of rodent-borne zoonoses transmission August 2020. The landfill is situated (-7.9591017 S; (Gholipoury et al., 2016). 112.6379437 E) near Malang city, East Java. Almost local Scientific literature associated to rodent-borne people work as scavengers and have a semi-permanent parasites in Indonesia insists on the role of rodents house around trapping locations. Urban rodents were as biological vector of ectoparasites (e.g., trapped around local community lives. The location was Xenopsylla cheopis, Hoplopleura pacifia, Poliplax closed with railroad and waterway (Figure 1). All spinulosa, and Ornithonyssus bacoti), zoonotic rodents were trapped alive using a rectangular metal endoparasites by helminths (e.g., Capillaria hepatica, trap baited with fried fish, salted fish, and roasted Gongylonema neoplasticum, Hymenolepis diminuta, coconut. Rodents capture were transported using a Hymenolepis nana, and Syphacia muris) and blood plastic bucket to the laboratory and euthanized protozoa particularly Trypanosomiasis and Leish- (ketamine:xylazine) with the guidelines for the maniasis (Afililla et al., 2017; Ernawati & Priyanto, humanely, following which, age, weight and sex 2013; Musyaffa et al., 2020; Sari et al., 2020). determined (AVMA, 2020; Tijjani et al., 2020b). Rodent not only spread infectious diseases but also made economic, and social problems. Animal Identification Nowadays, research in rat needs, as a first Identify the rodent species by morphological concern, to identifying most reservoirs of rodent- measurements and physical appearances. External borne zoonoses and mapping potential regions identification and inspection based on that occur of transmission, and minimizing measurements head and body length, weight, sex transmission risk in humans (Han et al., 2015). and maturity determination of development stage Considering the distribution of Rodent-borne consist of juvenile, sub-adult, and adult. It an help to zoonoses throughout Indonesia especially in correctly identify each animal to the species level urban area like Malang city, East Java. Preliminary based on characteristics (Herbreteau et al., 2011). study needed to provide local data at the landfill in Blimbing sub-district, Malang city. Based on Examination and Identifications of Ectoparasites information from the local community, in this area, there were wild rats that existed. The study Inspect and collection of ectoparasites from an aims to undertake the diversity of rodents anesthetized rat with a comb or dissecting forceps parasites as potential of zoonotic parasitic which to find ectoparasites in the fur. For better risks that threaten health. examination, the body of an anesthetized rat can take

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A B

Figure 1 The study area, in Blimbing sub-district, Malang City, East Java showing the study sites circled in red (A) and the local community lives in landfills area around the railroad (B).

head and underbody, fleas, and lice usually with Giemsa. The prepared slides were viewed predilection on the lower part of the abdomen, and under microscope Olympus CX-21 (Olympus Corp., underbody. For chigger mites usually in the ears Tokyo, Japan) with suitable magnification power part. Any ectoparasites were placed in a vessel tube (100x-1000x) and captured by Opti Lab Advanced containing 70% alcohol. Each tube was used for each Plus camera (Miconos Corp., Yogyakarta, Indonesia) rodent and labeling. After that, removed (Gholipoury et al., 2016). ectoparasites for processing used alcohol fixation After identification of parasites, the prepared increase concentration, clearing with xylene, and slides were measured by ImageJ software to know mounting on microscope slides for identification the length total in micrometers. Data were analyzed with standard protocols (Herbreteau et al., 2011; using descriptive study. Paramasvaran et al., 2009).

RESULTS AND DISCUSSION Examination and Identifications of Endoparasites and Blood Protozoan A total number of eight rodents including Rattus norvegicus (87.5%) and Suncus murinus (12.5%) were Rats were dissected following the protocol capture in this study. 50% of sampledf rodents were previously described (Herbreteau et al., 2011). Fecal male, 37.5% were female, and 12.5% of female the samples from gastrointestinal tracts (GITs) were musk shrew. Average weight of sample were male examined by wet smear after fecal flotation with 128,2g and 204.7g were female. Parasite infestation, saturated sodium chloride solution to detect including endoparasites was detected 100% and helminth eggs, larva, cyst, and parasite’s ova. Blood ectoparasites was detected in 87.5%. Total number collection by cardiac puncture. The blood smears endoparasites infection of rodents were 100%, must be used fresh blood added with anticoagulant infestation of ectoparasites were 87.5% and blood (EDTA). The thin and thick blood smears were stained protozoa were 37.5% (Table 1).

Three species of endoparasites (Helminths) were Most parasite infestation of wild rat was found in fecal samples from gastrointestinal tracts detected near semi-permanent house in Landfill (GITs) of wild rat and the musk shrew (Table 2). area. Syphacia obvelata also called pinworm was They were classified as Cestodes (one species) and collcted primarily in the the musk shrew. Nematoda (two species). Every species was Additionally, the recorded data showed high identification from wet smear after fecal flotation infections of Hymenolopis nana in all wild rat. and examine based on egg morphology (Baker, Rodents play a a major role in transmission of a 2007). After examined, all of them (Hymenolopis large number of infectious diseases to human. nana, Syphacia obvelata, and Nippostrongylus Immediately, Rodent-borne disease have multiple brasiliensis) have been categories as zoonotic effects on public health (Meerburg et al., 2009). We parasites (Figure 2). The protozoans were observed capture eight small mammals that included of using microscopic examination and in this study rodents and shrews. A total number of seven wild found Trypanosoma lewisi infection of three wild rat rat (Rattus norvegicus) identifications from urban (Figure 2). Two species of ectoparasites (flea and area and one species of the musk shrew. mite) were infestation in the fur. One species of Almost wildrats infected of ectoparasites, and fleas (Siphonaptera), Xenopsylla cheopis, on the endoparasites, this is indicated that the urban lower part of the abdomen, and the ventral body. habitat such as Blimbing sub-district has a potential The mites, Echinolaelap echidninus, found on the relationship with infestation of parasites. The host femur (Table 2). In most cases, many species of fleas (Rattus norvegicus) and the musk shrew (Suncus especially Xenopsylla cheopis are biological vector of murinus) of the majority of the endoparasites trypanosomes (Figure 2). related with medical importance. Most wild rats with

Table 1 Identification of rodent species and parasites infestation in Blimbing sub-district

Sex Age Parasite Family Species Female Male Juveniles Adult Ecto Endo Blood Muridae Rattus norvegicus 3(37.5%) 4 (50%) 1(12.5%) 6(75%) 7(87.5%) 7(87.5%) 3(37.5%) Soricidae Suncus murinus 1(12.5%) 0 1(12.5%) 0 0 1(12.5%) 0 Total 50% 50% 25% 75% 87.5% 100% 37.5%

Table 2 Percentage of parasites infestation in Blimbing sub-district

Present of Infested Parasite Species Characteristic Host samples (%) Endoparasites Cestodes Rn 87.5 (n=7) Hymenolopis nana Dwarf Tapeworm Sm 12.5 (n=1) Nematoda Syphacia obvelata Pinworm Sm 12.5 (n=1) Nippostrongylus brasiliensis Hookworm Rn 87.5 (n=7) Sm 12.5(n=1)

Ectoparasites Xenopsylla cheopis Rat flea Rn 50 (n=4) Echinolaelap echidninus Rat mite Rn 62.5 (n=5)

Blood protozoa Trypanosoma lewisi Hemoflagellates Rn 37.5 (n=3) Note: n= Total number of rodent infestations; Rn= Rattus norvegicus; Sm= Suncus murinus

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Figure 2 Nippostrongylus brasiliensis (A); Hymenolopis nana (B); Syphacia obvelata (C) identification from gastrointestinal tracts (GITs) with magnification 40x; Echinolaelap echidninus (D) ventral side showing chelicera (C), palpus (P), sternal plate (sp), epigynial plate (ep), leg segments [coxa (c), trochanter (t), femur (f), genu (g), tibia (b), tarsus (s) and pulvillus (v)]; Xenopsylla cheopis (E) showing Aedeagal apodeme of male (aa); Trypanosoma lewisi in the blood showing posterior margin(P), kinetoplast (K), nucleus (N), flagella (F). Scale-bar: 5 μm.

infection of endoparasites closely found in areas the caecum. In nematodes, the dominant with human activities. Every day, local community in preferences for Syphacia muris commonly infect this study area would be seen a wild rat wandering rodents (e.g., laboratory rats, wild rats, and mice) around the landfill and sewer. Present study (Khalil et al., 2014; Priyanto & Ningsih, 2014). reported that urban area and market town can Nippostrongylus brasiliensis is a trichostrongylid being a habitat for rodents and shrew (e.g., Rattus nematode in superfamily Trichosngyloidea r.diardii, Rattus tanezumi, Rattus norvegicus, and commonly found in small intestine. Pathological Suncus murinus (Dina & Ustiawan, 2013; Mohd Zain effect of the infections cause inflammation in the et al., 2012; Paramasvaran et al., 2009). skin, lungs, and intestine (Baker, 2007). Previous Rodents are well-known reservoirs and host study reported the prevalence of Nippostrongylus for number of infectious diseases. In this study, brasiliensis is 85.7% with parasites indexes 2-118 found endoparasites from GITs included individual parasites in each host, respectively Nippostrongylus brasiliensis, Hymenolopis nana, (Musyaffa et al., 2020). This is appropriate the high Syphacia obvelata (Figure 2). Recent study in risk of transmission of endoparasites to humans. Surabaya city, one of capital city in Indonesia In this work, total three Rattus norvegicus blood located near Malang city reported that prevalence samples were infected with Trypanosoma lewisi of Hymenolepis nana in 42% (Prasetyo, 2016). This (37.5%) (Figure 2). The prevalence of trypa- parasites as well-known of zoonotic agent in nosomiasis of wild rats in Banyuwangi city, East Java human lived in intestines, also called hymeno- was 1.67%. The infection of Trypanosoma lewisi in lepiasis caused by Hymenolepis diminuta, and wild rats has a potential risk to transmitted human Hymenolepis nana. Infections of dwarf tapeworm (Afililla et al., 2017). The distribution of the blood have been seen more often among children protozoa is worldwide. In Iran higher rates of (Anorital, 2014; Prasetyo, 2016). In this study, infection have been reported in Trypanosoma lewisi oxyurids (Syphacia muris) from shrew are found in in 11.54% (Seifollahi et al., 2016). In the current study,

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